The present invention relates to a very rapid method for the detection of polyamines, and to a kit for use in such method, in order to diagnose bacterial vaginosis.
In gynecological practice, bacterial vaginosis is one of the major reasons for visits by patients to physicians. Vaginosis associated with Gardnerella, may affect 5-40% of the female population attending medical practices or clinics. Anaerobic infections are accompanied by Gardnerella organisms, which become attached to desquamated vaginal cells. These cells are visible under the microscope as "clue cells". The present diagnosis and treatment of such infections is largely based on clinical symptoms and the microscopic appearance of the vaginal discharge with regard to clue cells. Further characteristics of bacterial vaginosis infections include elevated pH values, a homogenous malodorous vaginal discharge, and the production of a foul odor when potassium hydroxide is added to the discharge (the "whiff" test). Clinical suspicion may be confirmed by elaborate and time consuming microscopic and culturing techniques. Interpretation of the results in the existing methods of diagnosis, and the laboratory techniques involved, require considerable skills. Moreover, Gardnerella cultures do not correspond to the clinical syndrome.
The following is a list of relevant background references.
1. Amsel, R., Totten, P. A., et al, "Non-specific Vaginitis Diagnostic Criteria and Microbial and Epidemiological Associations", Amer. J. Med., 74: 14 (1983). PA1 2. Chen, K. C. S., Amsel, R., Eschenbach, D. A., Holmes, K. K., "Biochemical Diagnosis of Vaginitis: Determination of Diamines in Vaginal Fluids", J. Infect. Dis., 145: 337-345 (1982). PA1 3. Gardner, H. L., Dukes, C. D., "Haemophilus Vaginalis Vaginitis: A Newly-Defined Infection previously Classified as Non-Specific Vaginitis", Am. J. Obstet. & Gynaecol., 69: 962-976 (1955). PA1 4. Clarke Secor, R. Mimi, "Bacterial Vaginosis: A Comprehensive Review", Nursing Clinics of North America, 23(4): 865 (1988). PA1 5. Nelson, M. S., "Clinical Diagnosis of Bacterial Vaginosis", Amer. J. of Emergency Medicine, 5: 488 (1987). PA1 6. Sehgal, S. C., Nalini, V., "Role and prevalence of Gardnerella Vaginitis in Anaerobic Vaginosis", Infection, 18(2): 83/25 to 85/27 (1990). PA1 7. Livengood, C. H. III, Thomason, J. L., Hill, G. B., "Bacterial Vaginosis: Treatment with Topical Intravaginal Clindamycin phosphate", Obstet. & Gynaecol., 76(1): 118-123. PA1 8. Spiegel, C. A., Amsel, R., Holmes, K. K., "Diagnosis of Bacterial Vaginosis by Direct Gram Stain of Vaginal Fluid", J. of Clinical Microb., 1983 (July): 170-177. PA1 9. Thomason, J. L., Gelbart, S. M., Anderson, R. J., Walt, A. K., Osypowski, P. J., Broekhuizen, F. F., "Statistical Evaluation of Diagnostic Criteria for Bacterial Vaginosis", Am. J. Obstet. & Gynaecol., 162(1): 155-160 (1990). PA1 10. Hillier, S., Krohn, M. A., Watts, D. H., Wolner-Hanssen, P., Eschenbach, D., "Microbiological Efficacy of Intravaginal Clindamycin Cream for the Treatment of Bacterial Vaginosis", Obstet. & Gynaecol., 76(3): 407-413 (1990). PA1 11. Eschenbach, D. A., Hillier, S., Critchlow, C., Stevens, C., DeRouen, T., Holmes, K. K., "Diagnosis and Clinical Manifestations of Bacterial Vaginosis", Am. J. Obstet. & Gynaecol., 158(4): 819-828 (1988). PA1 (a) contacting a sample suspected of containing polyamine-associated bacteria with a liquid which comprises a mixture of sodium chloride and water; PA1 (b) mixing the thus-contacted liquid with a mixture of indigo carmine, sodium carbonate (preferably anhydrous) and dimethyl sulfoxide; PA1 (c) observing whether a blue color develops in five minutes indicating the presence of polyamines; PA1 (d) in the case that a blue color develops in step (c), visually comparing its intensity with a predetermined scale for estimating the presence and concentration of polyamines (a green coloration is considered to be clinically insignificant, a brown coloration indicates excessive amounts of polyamines and the sample must be diluted and the test repeated); PA1 (e) spectrophotometrically measuring the relative proportions of blue and green colorations in the test solution, in order to quantify the amount of polyamines present. PA1 (1) a finely-ground admixture of indigo carmine and sodium carbonate; PA1 (2) a sodium chloride solution; PA1 (3) dimethyl sulfoxide; and as optional additional components, at least one of the following components (4), (5) and (6), namely: PA1 (4) a color scale for enabling the presence and concentration of polyamines to be estimated by comparing with said scale the intensity of the blue color which may be obtained by contacting a test specimen suspected of containing such polyamines with components (1), (2) and (3); PA1 (5) a sterile swab for collecting said test specimen; and PA1 (6) a test vessel adapted for mixing components (1), (2) and (3), and said test specimen.